Modulation of the S100A9/CD33 pathway by the flavonoids ICA and ICT on the tumor microenvironment Background: Myeloid-derived suppressive cells are a recently derived component of the tumor microenvironment that has been determined to have a pivotal role in the maintenance and progression of cancer. These cells contribute to the upregulation of inflammatory correlated with tumor progression such as MRP8/9. Flavonoids are a group of polyphenolic compounds, commonly found on plants with many biological abilities that could prove useful in the battle against disease including anti-inflammatory, antioxidant and, more recently, anti-tumorigenic properties. Those obtained from Herba Epimedii, specifically its active compound icariin (ICA) and a derivative named 3, 5, 7-Trihydroxy-4'-methoxy-8-(3-hydroxy-3- methylbutyl)-flavone (ICT), have been shown to have strong immunomodulatory as well as anti-tumorigenic effects. Hypothesis and objective: We have recently shown that both of these compounds have potent modulatory effects that involve the downregulation of MDSC and MRP8/14 from the tumor microenvironment as well as the deactivation of STAT3, NF-kB and MAPK pathway components, cytotoxicity of tumor cells in vitro and the reduction of tumor burden in a mice model of cancer. While strides have been made in recognizing the role of these compounds on their therapeutic potential against cancer, their mechanisms of action is poorly understood. In particular we hypothesize that the effect on MDSC is linked to the inhibition of PDE5 just as seen with synthetic drugs like Viagra. We also hypothesize that the effect of these compounds on inflammatory pathways is mediated by the activation and perhaps upregulation of phosphatases, in particular PP2A as has been suggested previously. This particular effect might be linked to the direct cytotoxic effect of thes compounds on cancer cells. It is also possible that in vivo these drugs could have a direct impact on the architecture of the tumor microenvironment that lead to its destabilization and the inhibition of tumor growth. Therefore the study of the mechanisms involved in the modulation of MDSC and inflammatory markers linked to cancer by ICA and ICT would provide the insight to bring these compounds one step closer to the clinic for future therapeutical applications. Specific aims: Aim1: To understand the role of PDE5 inhibition on ICA/ICT action. Aim 2: To investigate the role of phosphatases on the deactivation of pro-inflammatory mediators by ICA and ICT. Aim 3: To study the regulatory mechanisms of ICA/ICT on the in vivo tumor microenvironment. Study design: For aim 1, we will first continue in silico modeling that demonstrates the specificities of the binding of ICT with PDE5. We will continue our assessment of this binding with mechanistic in vitro studies and determine if the mechanism is similar to that of other PDE5 inhibitors like sildenafil, our control. We will also determine if some of the effect of PDE5 inhibition affect tumor cells as well. For aim 2, we will determine the role of phosphatases on the deactivation of key pro-inflammatory pathways NF-kB, Akt, ERK, p38, and Stat3. Are PP2A and SHP1 the only phosphatases? Is there a specific type of phosphatases that gets activated by ICA and ICT. For aim 3, we will do in vivo studies to understand the role of these pathways and the importance each one of them plays in the modulatory role of ICA and ICT.